The long-term goal is to compare the different mechanisms by which HSV1 and HSV2 establish latent infection and reactivate to cause recurrent disease, particularly the involvement of the autonomic nervous system (ANS) pathways in these processes. HSV1 and HSV2 have different patterns of latent infection and different patterns of recurrent disease. While autonomic neurons have been recognized as a site for HSV infection for many years, their contribution to recurrent viral disease has not been closely examined, and differences in autonomic infection may contribute to the clinical and molecular patterns that differentiate HSV1 and HSV2. The central hypothesis is that HSV1 and HSV2 are capable of differentially establishing latency in, and reactivating from, specific types of autonomic neurons to cause some portion of recurrent HSV disease, regulated in part by the latency-associated transcript (LAT). The objectives of this proposal are 1) to compare the preferential pathways by which HSV1 and HSV2 reach the central nervous system (sensory or autonomic pathways), by using biologically relevant primary neuronal culture Campenot chamber infection systems in vitro and a guinea pig infection model (both genital and ocular) in vivo; 2) to examine the capabilities of HSV1 and HSV2 to reactivate from autonomic neurons, using a Campenot chamber infection and reactivation model in vitro, by reactivating autonomic neurons from latently infected mice ex vivo, and by using a chemical sympathectomy model in vivo to determine if pathway ablation alters infection and reactivation patterns; and 3) to demonstrate that the latency-associated transcript (LAT) region of the HSV genome regulates autonomic neuron specificity for HSV1 and HSV2 productive infection, by using chimeric viruses in which the LAT region has been swapped between HSV1 and HSV2 in in vitro and in vivo infection models to determine if switching LAT regions also switches patterns of infection and reactivation, as well as using protein-DNA binding assays to determine the host or viral proteins that differentially interact with the LAT regions of HSV1 and HSV2. Results from these studies will contribute to the overall understanding of HSV pathogenesis and suggest auxiliary treatments to prevent reactivation specifically from autonomic neurons to improve clinical outcomes of HSV-related recurrent disease. Completion of these studies will also provide the basic foundation for future R01 funding.